The ability to detect both L-homocysteine and L-methionine is critical for accurate screening for homocystinuria. This research project, if successful, will result in a selective, quantitative assay for L-homocysteine and L- methionine in biological fluids based on the reaction catalyzed by methionine gamma-lyase. The enzyme which is highly selective for L- methionine and L-homocysteine will be isolated from a strain of Pseudomonas and characterized. By coupling the enzymatic decomposition of L-methionine or L-homocysteine to an NADH-oxidizing dehydrogenase, rapid spectrophotometric read-out will be achieved. Conditions for quantitation of L-methionine and L-homocysteine will be developed, and the upper and lower limits of sensitivity will be determined. Possible interference by components of biological fluids will be assessed. Finally, methionine gamma-lyase will be immobilized and tested to establish the feasibility of a test strip for rapid, routine measurements. The commercial application for this work is immediate as a replacement for the Guthrie microbiological assay in neonatal screening, which is unable to detect L- homocysteine. Improved early diagnosis of homocysteinuria will permit more timely and effective treatment, resulting in lower costs and dramatically improved outcomes for homocystinurics. Further opportunities exist for development of a rapid test strip. PROPOSED COMMERCIAL APPLICATIONS: Neonatal screening assay for homocystinuria; home and doctor's office test strip for measurement of L-methionine and L-homocysteine concentrations in urine or blood.